Acoustophoresis device having improved dimensions

ABSTRACT

Systems and methods for cleansing blood are disclosed herein. The methods include acoustically separating target particles from elements of whole blood. The whole blood and capture particles are flowed through a microfluidic separation channel formed in a thermoplastic. At least one bulk acoustic transducer is attached to the microfluidic separation channel. A standing acoustic wave, imparted on the channel and its contents by the bulk acoustic transducer, drives the formed elements of the blood and target particles to specific aggregation axes.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims priority to U.S. Provisional Patent ApplicationNo. 62/519,630 filed on Jun. 14, 2017, which is herein incorporated byreference in its entirety.

BACKGROUND OF THE DISCLOSURE

Sepsis is a disease with a very significant public health impact thathas stubbornly resisted new therapies. Antibiotics are the only realtherapeutic option, yet sepsis can be caused by over 100 bacteria andmany fungi, so a universal antibiotic is not a realistic option; theantibiotics and antifungals used have significant complications and areoften unsuitable for fragile patients. The concept of cleansing theblood has been tried previously without success. Previous bloodcleansing concepts have included laboratory scale methods ofcentrifugation, capillary electrophoresis, liquid chromatography, fieldflow fractionation, and liquid-liquid extraction. These devices havefailed to deliver continuous flow cleansing devices. In addition tooften discarding large portions of the blood, current cleansing devicesmay rely on diluents, sheath flow, controlled solution conductivity,costly microfabricated on-chip materials, and toxic additives.

SUMMARY OF THE DISCLOSURE

The present disclosure described new design rules for plastic-baseddevices. Specifically, the disclosure describes ratios between operationfrequency, wall thickness, and channel width that can be used tocalculate optimized channel dimensions for separation devicesconstructed in plastic.

Silicon, glass, or metal devices are commonly used for acoustophoresisbecause the rigid channel walls provide a near ideal acoustic boundaryagainst the sample fluid, enhancing the required standing waveresonance. This ideal boundary simplifies design because the resultantreduction in mathematical complexity allows for the development ofmodels that can be used to calculate the resonant modes in thechannel-fluid system. However, the rigid materials previously used areexpensive and slow to manufacture, have poor compatibility with manybiological samples, and are unlikely to be acceptable for massproduction of disposable laboratory tools.

According to at least one aspect of the disclosure, a cleansing devicecan include a first thermoplastic substrate. The substrate can include aseparation channel having a width, a first wall, a second wall, and afloor. The floor can be configured to couple with an acoustic transducerand a thickness of the first wall and the second wall can be based onthe width of the separation channel. The device can include a firstinlet configured to introduce a fluid into a proximal end portion of theseparation channel. The device can include a first outlet at thedownstream portion of the separation channel positioned substantiallyalong the longitudinal axis of the separation channel. The device caninclude a second outlet at the downstream portion positioned adjacent afirst wall of the separation channel. The device can include a secondthermoplastic substrate coupled with the first thermoplastic substrateand defining a roof of the separation channel.

In some implementations, the thickness of the floor can be based on thewidth of the separation channel. The width of the separation channel canbe between 0.1 mm and 3 mm. The thickness of the first wall and secondwall can be based on a velocity of an acoustic wave through the fluid.The thickness of the first wall and the second wall can be based on avelocity of the acoustic wave through the first thermoplastic substrate.The height of the separation channel can be based on the width of theseparation channel.

In some implementations, a ratio of the separation channel to the heightof the separation channel can be between 2 and 2.5. The floor of theseparation channel can have a first thickness and the roof of theseparation channel can have a second thickness. The first thickness canbe greater than the second thickness. A ratio of the thickness of thefloor of the separation channel to a width of the separation channel canbe between about 0.5 and 1.

According to at least one aspect of the disclosure, a method to cleansefluid can include providing a separation device. The device can includea first thermoplastic substrate that can include a separation channelhaving a width, a first wall, a second wall, and a floor. The floor canbe configured to couple with an acoustic transducer and a thickness ofthe first wall and the second wall can be based on the width of theseparation channel. The device can include a first inlet configured tointroduce a fluid into a proximal end portion of the separation channel.The device can include a first outlet at the downstream portion of theseparation channel positioned substantially along the longitudinal axisof the separation channel. The device can include a second outlet at thedownstream portion positioned adjacent a first wall of the separationchannel. The device can include a second thermoplastic substrate coupledwith the first thermoplastic substrate and defining a roof of theseparation channel. The method can include flowing a fluid through thefirst inlet. The fluid can include undesirable particles. The method caninclude driving, with a standing acoustic wave generated by the acoustictransducer, the undesirable particles toward the first wall of theseparation channel.

In some implementations, the method can include applying the standingacoustic wave through the floor of the separation channel. The width ofthe separation channel can be between 0.1 mm and 3 mm. The thickness ofthe first wall and second wall can be based on a velocity of theacoustic wave through the fluid and a velocity of the acoustic wavethrough the first wall and the second wall. The thickness of the firstwall and the second wall can be based on a velocity of the acoustic wavethrough the first thermoplastic substrate. The height of the separationchannel can be based on the width of the separation channel.

In some implementations, a ratio of the separation channel to the heightof the separation channel can be between 2 and 2.5. The floor of theseparation channel can have a first thickness and the roof of theseparation channel can have a second thickness. The first thickness canbe greater than the second thickness. A ratio of the thickness of thefloor of the separation channel to a width of the separation channel canbe between about 0.5 and 1.

The present disclosure discusses design rules for the manufacture ofseparation channels in plastic, including thermoplastics and other lossyplastics. While acoustic separation devices can be constructed inplastic using the same design rules for silicon, glass, or metaldevices, the design rules do not provide optimized, plastic-baseddevices because the simplified analysis no longer applies since thechannel walls can no longer be considered ideally rigid.

BRIEF DESCRIPTION OF THE DRAWINGS

The skilled artisan will understand that the FIGS., described herein,are for illustration purposes only. It is to be understood that in someinstances various aspects of the described implementations may be shownexaggerated or enlarged to facilitate an understanding of the describedimplementations. In the drawings, like reference characters generallyrefer to like features, functionally similar and/or structurally similarelements throughout the various drawings. The drawings are notnecessarily to scale, emphasis instead being placed upon illustratingthe principles of the teachings. The drawings are not intended to limitthe scope of the present teachings in any way. The system and method maybe better understood from the following illustrative description withreference to the following drawings in which:

FIG. 1 is a block diagram of an example system for cleansing blood.

FIG. 2 is a top view of an example single-stage separation channel, suchas can be used in the system of FIG. 1.

FIG. 3 illustrates a cross sectional view through section A illustratedin FIG. 2.

FIG. 4A is a cross sectional view of an example single-stage separationdevice, such as the separation device of FIG. 2, mounted to a bulktransducer.

FIG. 4B illustrates a top view of an example separation device withmultiple separation channels.

FIG. 4C illustrates a block diagram of an example method to design aseparation device.

FIG. 5A is a cross sectional view of an example single-stage separationchannel, as depicted in FIG. 2, containing a plurality of particleslacking an active acoustic transducer.

FIG. 5B is a cross sectional of an example single-stage separationchannel, as depicted in FIG. 2, containing a plurality of particlesadjacent to an active acoustic transducer.

FIG. 6A is a top view of a separation channel, as depicted in FIG. 2, inwhich fluid is flown through the channel without the application of thestanding acoustic wave.

FIG. 6B is a top view of a separation channel, as depicted in FIG. 2,after the application of a standing acoustic wave.

FIG. 7 is a cut away of an example lipid-based capture particle.

FIGS. 8A-8E are illustrations of the components and use for a captureparticle, as depicted in FIG. 7.

FIG. 9 is a flow chart of an example method for cleansing blood with asingle-stage separation channel, as depicted in FIG. 2.

FIGS. 10A-10C illustrate plots of the performance of separation devicesat several flow rates using the design rules described herein andbaseline devices, in terms of each device's ability to focus red bloodcells to a central aggregation axis.

FIGS. 11A-11C illustrate plots of the performance of the experimentaldevice against the baseline in terms of each device's ability to focusred blood cells to a central aggregation axis and out a center outletport.

FIG. 12A illustrates the relative performance of the baseline device asit compares to the experimental design in terms of flow rate with powerheld constant at 1 W.

FIG. 12B illustrates the performance of the experimental design andbaseline design when holding flow rate at 50 μL/min while varying power.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

The various concepts introduced above and discussed in greater detailbelow may be implemented in any of numerous ways, as the describedconcepts are not limited to any particular manner of implementation.Examples of specific implementations and applications are providedprimarily for illustrative purposes.

The present system and methods described herein generally relate to asystem for cleansing blood. Accordingly, in various implementations, thedisclosure relates to the acoustically separating particles from theblood (or other fluids) via high throughput microfluidic arrays. Theseparated particles can generally be referred to as target particles.The target particles can be undesirable particles in cases where thedevices described herein are used for purification or filtering. Inother implementations, the target particles can be cells or particlesthat are removed from a fluid for further study or processing (e.g., thetarget particles are desired particles). For example, the targetparticles can be cells that are removed from a fluid such that adiagnostic analysis can be performed on the cells. In certainimplementations, in part to overcome the prior deficiencies with thepoor performance of acoustic separation on small particles, prior toacoustic separation of the blood, capture particles are introduced andmixed with the blood to form complexes with the undesirable particles,yielding particles large enough to be effectively and efficientlytargeted by acoustic separation.

FIG. 1 illustrates a system 100 for cleansing blood by removing wastematerial such as bacteria, viruses and toxins. In the system 100, bloodis removed from a patient via an intravenous line 102. The blood is thenpumped to a mixing chamber 104 by a first pump 103. In the mixingchamber 104, capture particles are mixed with whole blood. Thecomponents of the capture particles are stored in a reservoir 107. Fromthe reservoir 107, the capture particles are pumped by a second pump 106into the mixing chamber. The capture particles are formed as thecontents of the reservoir 107 are extruded from a micronozzle 105 at theentrance to the mixing chamber 104. From the mixing chamber 104, thewhole blood and capture particles enter a manifold system 107. Themanifold system 107 distributes the whole blood and capture particles toa plurality of separation channels contained within the microfluidicflow chamber 108. The microfluidic flow chamber 108 sits atop at leastone bulk piezoelectric acoustic transducer 109. The acoustic wavesgenerated by the bulk piezoelectric acoustic transducers are used tofunnel the contents of the whole blood and capture particles to specificoutlets of the separation channels. As the whole blood flows through themicrofluidic flow chamber 108, cleansed blood flows to a first outlet110. After exiting the first outlet 100, the cleansed blood returns tothe patient 101 via a second intravenous line 111. The capture particlesand other waste material removed from the blood exit the microfluidicflow chamber 108 via a second outlet 112. Next, the waste material andcapture particles enter a waste collection unit 113. In the wastecollection unit 113, the capture particles are separated from the wastematerial. Once separated, the waste material is discarded, and thecapture particles are returned to the reservoir 107 by tubing 114. Oncereturned to the reservoir 107, the capture particles are reused in thesystem to remove additional waste material from whole blood as itcontinues to flow through the system.

The system 100, as illustrated, includes a pump 103 for moving bloodfrom the patient 101 to the mixing chamber 104. In some implementations,the pump operates continuously, while in other implementations the pumpworks intermittently and only activates when the level of whole blood inthe mixing chamber 104 or manifold falls below a set threshold. In someimplementations, the flow rate of the pump is configurable, such thatthe rate the blood exits the patient can be configured to be faster orslower than if no pump was used. In yet other implementations, noexternal pump is required. In this example, the blood is transported tothe mixing chamber 104 by the pressure generated by the patient's ownheart. In some implementations, the patient 101 is connected to a bloodpressure monitor, which in turn controls the pump. Example pumps caninclude, but are not limited, to peristaltic pumps or any other pumpsuitable for flowing blood.

As illustrated in the system 100, capture particles are also pumped intothe mixing chamber. A second pump 106 pumps the ingredients to form thecapture particles from a reservoir 107 to the mixing chamber 104. Insome implementations, the components of the capture particles arecontinuously agitated in the reservoir 107 in order to keep thecomponents well mixed. The components are formed into capture particlesas they enter the mixing chamber 104. The components enter the mixingchamber 104 through a micronozzle 105. In some implementations, themicronozzle 105 injects the capture particles into the mixing chamber104. In other implementations, the micronozzle 105 injects the captureparticles into the manifold system 107, and in yet other implementationsthe micronozzle 105 is positioned such that it injects capture particlesdirectly into the separation channels of the microfluidic chamber 108.In some implementations, the micronozzle 105 is a micro-machined nozzle,configured to allow a specific amount of the capture particle componentsthrough the nozzle at a given time. In some implementations, themicronozzle is an array of micronozzles. In yet other implementations,the micronozzle is a membrane with pores. The pump 106 is configured toflow the contents of the reservoir through the micronozzle 105 at apredetermined rate such that the amphipathic characteristics of themolecules of the components of the captures particles cause the captureparticles to spontaneously form as they exit the micronozzle 105.

In some implementations, a micronozzle is not used to generate thecapture particles. In these implementations, the capture particles arepremade. The capture particles are then stored in the reservoir andintroduced into the system by the pump 106 at either the mixing chamber104, manifold system 107, and/or the separation channels of themicrofluidic flow chamber 108. In some implementations, captureparticles are not used. The particles of the fluid can be driven to anaggregations axis based on the particle's inherent acoustic contrastfactor with respect to the fluid.

As illustrated in system 100, the whole blood containing targetparticles and the capture particles enter the mixing chamber 104. Insome implementations, the contents of the mixing chamber arecontinuously agitated to improve distribution of the capture particlesthroughout the whole blood and target particles such that the captureparticles bind to the target particles. In some implementations,anticoagulants or blood thinners are introduced into the mixing chamber104 to assist the blood as it flows through the system 100. In someimplementations, the mixing chamber 104 contains a heating element forwarming the contents of the mixing chamber 104.

The contents of the mixing chamber 104 then flow into the manifoldsystem 107, as illustrated by system 100. The manifold system 107 flowsthe whole blood, target particles, and capture particles into the inletsof the plurality of separation channels of the microfluidic flow chamber108.

In the illustrated system 100, the microfluidic flow chamber 108contains a plurality of separation channels. The capture particles andtarget particles are driven with standing acoustic waves to outlets. Insome implementations, the separation occurs during a single stage, whilein other implementations, the separation occurs over a plurality ofstages. In some implementations, the microfluidic flow chamber isdisposable.

As shown in the illustrations of system 100, the microfluidic flowchamber 108 sits atop a bulk piezoelectric acoustic transducer 109. Insome implementations, the system 100 contains a single bulkpiezoelectric acoustic transducer 109, while in other implementationsthe system 100 contains a plurality of bulk piezoelectric acoustictransducers 109.

In some implementations, the bulk piezoelectric acoustic transducer 109is glued to the microfluidic flow chamber 108. In other implementations,the microfluidic flow chamber 108 is clamped to the bulk piezoelectricacoustic transducer 109 so the microfluidic flow chamber may easily beremoved from the system. In other implementations, the adhesive materialconnecting the bulk piezoelectric acoustic transducer 109 to themicrofluidic flow chamber 108 is removable, for example by heating theadhesive.

The bulk piezoelectric acoustic transducer 109 imposes a standingacoustic wave on the separation channels of the microfluidic flowchamber 108 transverse to the flow of the fluid within the microfluidicflow chamber 108. The standing acoustic waves are used to drive fluidconstituents towards or away from the walls of the separation channelsor other aggregation axes.

More particularly, the dimensions of the separation channels areselected based on the wavelength of the imposed standing wave such thata pressure node exists at about the center or other interior axis of theseparating channel, while antinodes exist at about the walls of theseparation channel. The dimensions are discussed in relation to FIG. 4A.Particles are driven to different positions within the channel based onthe sign of their acoustic contrast factor at a rate which isproportional to the magnitude of their contrast factor. Particles with apositive contrast factor (e.g. the formed elements of blood) are driventowards the pressure node within the interior of the separation channel.In contrast, particles with a negative contrast factor are driven towardthe pressure antinodes. These principles are depicted and describedfurther in relation to FIGS. 5A and 5B.

Based on these principles, formed elements of blood can be separatedfrom capture particles and the target particles bound to the captureparticles. In one way, as described further in relation to FIGS. 2 and10, capture particles are selected to have negative contrast factors,which is opposite to the positive contrast factors of the formedelements of blood. Thus, in response to the standing acoustic wave, theformed elements are driven towards the resulting pressure node while thecapture particles are driven towards the antinodes.

This technique can be used in a single-stage separation system, asillustrated in FIG. 2. As whole blood and target particles (and in someimplementations, capture particles) mix in the mixing chamber 104 andcontinue to mix as flowing through the manifold system 107, the targetparticles enter the area of the separation channel where the standingacoustic wave is imparted. The standing acoustic wave drives the boundtarget particles to a specific axis (e.g., against the wall of theseparation channel) and the formed elements of the whole blood to asecond axis (e.g., the middle of the separation channel). Thus, thetarget particles can be collected from the edges of the separationchannel and disposed of while the cleaned blood is collected andreturned to the patient.

As illustrated in the system 100, the cleansed blood exits themicrofluidic flow chamber 108 at a first outlet 110. From there theblood is returned to the patient 101 via an intravenous supply line 111.In some implementations, the blood in the supply line 111 is reheated tobody temperature before returning to the patient 101. In otherimplementations, an infusion pump is used to return the blood to thepatient 101, while in the system 100 the pressure generated in thesystem by pumps 103 and 106 is adequate to force the blood to return tothe patient 101.

As illustrated in the system 100, waste material (e.g. the captureparticle and target particles) exits the microfluidic flow chamber 108and enters a waste collection unit 113. In some implementations, thewaste collection unit 113 contains a capture particle recycler. Thecapture particle recycler unbinds the target particles from the captureparticles. The capture particles are then returned to the reservoir 107via tubing 114. The target particles are then disposed of. In someimplementations, the target particles are saved for further testing.

While the system 100 is described above for the in-line cleansing of apatient's blood, in alternative implementations, the system 100 can beused to cleanse stored blood. For example, the system 100 can be used tocleanse collected blood for later infusion to help ensure the safety ofthe blood.

FIG. 2 illustrates an example single-stage separation channel suitablefor use within the microfluidic flow chamber 108 of the blood cleansingsystem 100. The separation channel includes an inlet 202, a flow channel203, a first outlet 204, a first outlet channel 206, a second outletchannel 207, and a second outlet 205. The separation channel ismanufactured in a sheet of material 201.

In FIG. 2, whole blood, target particles, and capture particles enterthe separation channel at the inlet 202 from the manifold system 107.The whole blood, target particles, and capture particles then flow thelength of the flow channel 203. The flow channel is subdivided intothree regions: an upstream region, a downstream region, and a migrationregion. The migration region lies between the upstream and downstreamregions, and is the region of the flow channel where the standingacoustic wave is imparted transverse to the flow of particles. As theformed elements of the whole blood, capture particles, and the targetparticles enter the migration region, the standing acoustic wave drivesthe capture particles bound to the target particles to the side walls ofthe separation channel, and the formed elements of the whole blood tothe center of the channel. The formed elements of the whole blood thenexit the separation channel through the outlet 204 located at about thecentral axis of the separation channel. The capture particles and targetparticles then exit the separation channel through the first and secondoutlet channels 206 and 207 which terminate in the second outlet 205. Insome implementations, the formed elements are driven to the walls of theseparation channel and the capture and target particles remain in thecenter of the separation channel.

In some implementations, the separation channel 200 can separate targetparticles from any fluid. As discussed above and later in relation toFIGS. 5 and 7, the separation channel 200 can be used to remove targetparticles from any fluid, so long as the characteristics of the captureparticle are appropriately selected. For example, selecting anencapsulated fluid such that its density and bulk modulus gives thecapture particle a contrast factor that distinguishes it from the fluidand other particles in the fluid. For example, the separation channel200 may be used to remove target particles from, but not limited to,blood plasma, blood serum, water, liquid food products (e.g., milk),lymph, urine, sputum, and cell culture media.

In the implementation of FIG. 2, the outlet 205 is formed from themerging of two outlet channels 206 and 207. In some implementations, thestreams do not rejoin but lead to separate outlet terminals.

In FIG. 2, the particles are separated in the same plane as the sheet ofmaterial 201 (i.e. particles are aligned to the left, right, or centerof the channel); however, in other implementations, the particles areseparated out of plane. For example, in some implementations, theparticles are aligned with the top, middle, or bottom of the channel.

In FIG. 2, the sheet of material 201 can include thermoplastics or otherlossy plastics, such as, but not limited to, polystyrene, acrylic(polymethylmethacrylate), polysulfone, polycarbonate, polyethylene,polypropylene, cyclic olefin copolymer, silicone, liquid crystalpolymer, polyimide, polyetherimide, and polyvinylidene fluoride. Thechannel can be manufactured by a number of manufacturing techniques,including, but not limited to, milling, molding, embossing, and etching.

FIG. 3 illustrates a cross sectional view 300 through section Aillustrated in FIG. 2. The wave 302 indicates the pressure amplitudeP(t) in a standing wave across the separation channel 200. The solidline and dashed line are alternate phases of the wave 302. Particlesaccumulate at the pressure node 304 where time averaged P is zero. Thefrequency and relative dimensions of fluid and walls can be selected asfractions of the acoustic wavelength.

FIG. 4A is an illustrative cross-section of a separation device 400similar to the separation channel depicted in FIG. 2, for example. Theseparation device 400 includes a top layer 401 sitting atop a bottomlayer 402. A separation channel 403 (which can also be referred to as alumen 403) is formed in the top layer 401. In some implementations, theseparation channel 403 is formed in the bottom layer 402 or acombination of the top layer 401 and the bottom layer 402. The bottomlayer 402 can form one wall of the separation channel 403, such as thefloor of the separation channel 403. The floor of the separation device400 is coupled with a bulk piezoelectric transducer 404. The wall orportion of material separating the separation channel 403 from thetransducer 404 can be referred to as the “floor” and does not have to bethe bottom wall of the separation channel. The “ceiling” is the wallopposite the floor. For example, if the transducer were coupled with thetop layer 401, the floor would form the top wall (as illustrated in FIG.4A) of the separation channel 403 and the ceiling would form the bottomwall of the separation channel 403. The separation device 400 is securedto the bulk transducer 404, by a coupling adhesive 405 and/or mechanicalclamp. In some implementations, the coupling adhesive is cyanoacrylateglue. In some implementations, the bulk piezoelectric transducer 404 canbe operated at a frequency between about 0.5 MHz and about 2.5 MHz, orbetween about 0.5 MHz and about 1 MHz.

The bottom layer 402 and top layer 401 of the separation device 400 aremanufactured from a substrate sheet. The substrate sheet can be made of,without limitation, one of the above described plastics. In someimplementations, the bottom layer 402 is manufactured by milling,embossing, molding, and/or etching. After creating the two layers, theycan be joined together by thermocompression, mechanical clamping,adhesive bonding, and/or plasma bonding. The separation device can sitatop the acoustic bulk transducer 404 such that the transducer 404 isseparated from the separation channel 403 by a distance of h_(s1). Forexample, the separation channel 403 can have a floor with a thickness ofh_(s1). In some implementations, the transducer 404 may be mounted to asidewall of the separation device 400 such that the transducer 404 isseparated from the separation channel 403 by a distance of w_(s). Thetransducer 404 imparts a standing acoustic wave of a specific wavelengththrough the bottom layer 402, separation channel 403, and top layer 401.The dimensions of the bottom layer 402, top layer 401, and separationchannel 403 are dependent on the selected wavelength.

In some implementations, the separation device 400 can include aplurality of separation channels 403. FIG. 4B illustrates an exampleseparation device 400 that includes a plurality of separation channels403. The separation device 400 can operate similar to a plurality of theseparation channels illustrated in FIG. 2 linked together in parallel.The outputs of the separation channels can be coupled together via themanifold 107. The manifold 107 can include a first outlet for thecollection of waste (or other fluid) and a second outlet for thecollection of cleansed or filtered fluid. Each of the separationchannels 403 can be separated from neighboring separation channels 403by air gaps 410. The air gaps 410 can be voids in the substrates 402 and401 that run along at least a portion of the length of the separationchannel 403. The air gaps 410 can extend from the top of the topsubstrate 401 to the bottom of the bottom substrate 402.

The design for a separation device 400 can be based on the theory ofresonant cavities. The device 400 cross section has dimensions of widthand height for both the fluid-filled separation channel 403 and for thesolid walls that are formed by the top layer 401 and the bottom layer402 and define the separation channel 403. In some implementations, theacoustic force moves particles in the horizontal (e.g., the lateral orwidth) direction and towards a pressure node in, for example, the centerof the separation channel 403. In these implementations, the analystscan place emphasis on the width dimensions. The vertical (height)dimensions can be ignored in some implementations. In someimplementations, the separation channel is assumed to have a “hard wall”boundary. The hard-wall boundary can imply that a wave is approximatelyperfectly reflected at the boundary between fluid and solid so that thetime averaged pressure P(t) attains a maximum at the fluid-solidboundary. The result of this one-dimensional, hard-wall analysis is thatthe fluid channel width can be one-half of an acoustic wavelength in thefluid and that the solid wall width on each side should be one fourth ofan acoustic wavelength in the solid. In some implementations, multiplesof one quarter are used (e.g., ½, ¾, 1 . . . ). The hard wall assumptioncan be used to design separation channels, for example, in glass orsilicon.

The term “acoustic wavelength”, denoted by λ, is defined by the speed ofsound c (acoustic velocity) in the material and the operating frequencyω,

λ=c/ω  (1)

Therefore, the acoustic wavelength has a specific value only for aspecific property and an operating frequency, for example:

λ_(f) =c _(f)/ω  (2)

λ_(s) =c _(s)/ω  (3)

for the fluid and solid respectively. Since a device is ordinarilyactuated at a single frequency in both fluid and solid, the above twoequations can be combined as:

$\begin{matrix}{\frac{c_{f}}{\lambda_{f\;}} = \frac{c_{s}}{\lambda_{s}}} & (4)\end{matrix}$

Further manipulation using the dimensions shown in FIG. 4A leads to aratio of width of channel to width of wall, expressed as:

$\begin{matrix}{{\frac{w_{s}}{w_{f}} = {\frac{n}{2}\frac{c_{s}}{c_{f}}}}{{n = 1},2,3,\ldots}} & (5)\end{matrix}$

and this ratio should apply for in principle for any operatingfrequency. Table 1 lists examples of the resulting ratio of wall widthto channel width.

TABLE 1 Width of fluid channel and bounding walls and their ratios inseveral experiments. c fluid c solid Fluid Solid (m/s) (m/s) w_(f) (mm)w_(s) (mm) w_(s)/w_(f) n saline silicon 1500 8490 0.377 1.072 2.84 1saline silicon 1500 8490 0.377 2.147 5.69 2 saline polystyrene 1500 24000.430 1.050 2.44 3 saline polystyrene 1500 2400 0.550 0.850 1.55 1.9

However, devices made of thermoplastics such as polystyrene cannot beaccurately represented by the hard-wall approximation discussed above,and the simplified analysis does not provide appropriate design rules.Design rules for devices constructed in plastic are described below. Thedesign rules for thermoplastics can be different because they do notcall for a half wavelength in the fluid as:

$\begin{matrix}{w_{f} = {{\lambda_{f}/2} = \frac{c_{f}}{2\omega}}} & (6)\end{matrix}$

Instead, for separation device 400 using plastics, the channel width canbe defined using design rules and dimension ratios (with respect to thewall thickness, wave speed in the fluid and plastic, and/or theoperating frequency) specific to plastics. For example, the channelwidth can be defined as:

$\begin{matrix}{{w_{f} \cong {\lambda_{f}/4}} = \frac{c_{f}}{4\omega}} & (7)\end{matrix}$

which results in increased performance of driving particles to pressurenodes in separation channels 400 constructed in thermoplastics.

Table 2 compares the channel width as it relates to acoustic wavelengthin both silicon and polystyrene devices (e.g., a plastic device).

c fluid c solid ω w_(f) w_(s) λ_(f)/2 Fluid Solid (m/s) (m/s) (MHz) (mm)(mm) (mm) w_(f)/λ_(f) saline silicon 1500 8490 2 0.377 1.072 0.375 0.50saline silicon 1500 8490 2 0.377 2.147 0.375 0.50 saline poly- 1500 24001.05 0.430 1.050 0.765 0.28 styrene saline poly- 1500 2400 0.632 0.5500.850 1.187 0.23 styrene

As illustrated in Table 2, the ratio defining the channel width in asilicon device is 0.5. In contrast, for a thermoplastic device, theratio is between 0.28 and 0.23. The ratio can be between about 0.15 andabout 5, between about 0.15 and about 0.4, between about 0.2 and about0.3, or between about 0.23 and about 0.28.

In one example, the above ratio of a thermoplastic device provides achannel with a width of 0.55 mm and a height of 0.25 mm. In someimplementations, the ratio of width to height is:

$\begin{matrix}{h_{f} = \frac{w_{f}}{2.2}} & (8)\end{matrix}$

Additionally, for thermoplastic based devices:

$\begin{matrix}{h_{s\; 1} = \frac{w_{s}}{0.88}} & (9) \\{h_{s\; 2} = {w_{s}/1.2}} & (10)\end{matrix}$

In some implementations, the w_(s)/h_(s1) ratio can be between about 0.5and about 5.0, between about 0.6 and about 3.0, between about 0.7 andabout 2.0, or between about 0.8 and about 1.0. In some implementations,the w_(s)/h_(s2) ratio can be between about 0.5 and about 5, betweenabout 1.1 and about 2, or between about 1.20 and about 1.5.

FIG. 4C illustrates a block diagram of an example method 450 to design aseparation channel. The method 450 can include selecting a separationchannel width (step 452). The method 450 can include selecting a fluid(step 454). The method 450 can include selecting a wall width (step456). The method 450 can include selecting a channel height (step 458).The method 450 can include selecting a thickness (step 460). The method450 can include selecting an operating frequency (step 462).

As set forth above, the method 450 can include selecting a separationchannel width (step 452). Also referring to FIG. 4C, the method 450 caninclude selecting the width of the separation channel 403. The width ofthe separation channel 403 can be measured as the portion of theseparation channel 403 parallel with the transducer 404 and the heightof the separation channel 403 can be measured as the portion of theseparation channel 403 perpendicular with the transducer 404. The methodcan include selecting the width w_(f) of the separation channel 403. Insome implementations, the separation channel width w_(f) can be in therange of about 0.1 mm and about 5 mm, between about 0.1 mm and about 4mm, between about 0.1 mm and about 3 mm, between about 0.1 mm and about2 mm, or between about 0.1 mm and about 1 mm. The separation channelwidth can be selected first based on the desired fluid throughput of theseparation device 400.

The method 450 can include selecting a fluid flow (step 454). The method450 can include selecting which fluid to flow through the separationchannel 403. The fluid can include blood. The method 450 can includedetermining the velocity of an acoustic wave through the fluid (c_(f)).

The method 450 can include selecting a wall width (step 456). Alsoreferring to FIG. 4A, the wall width selected at step 456 can be thewall width w_(s). The wall width w_(s) can be the width of a wallperpendicular to the transducer 404. The wall width w_(s) can becalculated according to Equation 5, with n set to 1 or 2. As illustratedin Equation 5, the wall width w_(s) can be dependent on the separationchannel width (w_(f)) selected at step 452 and the velocity of anacoustic wave through the fluid (c_(f)) selected at step 454. The wallwidth w_(s) can be based on the velocity of an acoustic wave through thesubstrate material (c_(s)) (e.g., the velocity of an acoustic wavethrough the material of the substrate 402 and the substrate 401).

The method 450 can include selecting the channel height (step 458). Alsoreferring to FIG. 4A, the channel height h_(f) can be selected accordingto Equation 8, above. As illustrated in Equation 8, the channel heighth_(f) can be based on the separation channel width (w_(f)) selected atstep 452. A ratio of the separation channel to the height of theseparation channel can be between about 2 and about 3.5, between about 2and about 3, between about 2 and about 2.5, or about 2.2.

The method 450 can include selecting the thickness (step 460). Themethod 450 can include selecting the thickness h_(s1) (e.g., the floorof the separation channel 403) and the thicknesses h_(s2) (e.g., theceiling of the separation channel 403) illustrated in FIG. 4A. Thethickness h_(s1) can be calculated using Equation 9. The thicknessh_(s2) can be calculated using Equation 10. The thickness h_(s1) and thethicknesses h_(s2) can be calculated based on the wall width w_(s)determined in step 456. The thickness h_(s1) and the thicknesses h_(s2)can be different thicknesses. In some implementations, the thicknessh_(s1) and the thicknesses h_(s2) can be the same thicknesses. In someimplementations, the thickness h_(s1) of the material between thetransducer 404 and the separation channel 403 can be greater than thethickness h_(s2) of the substrate 401. For example, both the thicknessh_(s1) and the thicknesses h_(s2) can be calculated using Equation 9 orEquation 10. A ratio of the wall width w_(s) to the thickness h_(s1) canbetween about 0.2 and about 1.5, between about 0.5 and about 1, betweenabout 0.8 and about 1, or about 0.88. A ratio of the wall width w_(s) tothe thickness h_(s2) can between about 0.75 and about 2.5, between about1 and about 2, between about 1 and about 1.5, or about 1.2.

The method 450 can include selecting an operating frequency (step 460).Also referring to FIG. 4A, the operating frequency can be the frequencyat which the transducer 404 is operated. The operating frequency can becalculated using Equation 7. The operating frequency can be calculatedusing the velocity of an acoustic wave through the fluid (c_(f))selected at step 454 and the separation channel width w_(f) selected atstep 452. In some implementations, the wavelength of the operatingfrequency is between about 2 and about 8, between about 3 and about 6,between about 3 and about 5.5, or between about 3.3 and about 5 timesthe separation channel width (w_(f)).

FIGS. 5A and 5B are cross sectional views of particles suspended in afluid as they flow through a separation channel similar to theseparation channel 200. For FIGS. 5A and 5B, the flow of the fluid istransverse to the plane of the drawings. In some implementations, thefluid is whole blood, and the particles are the formed elements andcapture particles. For illustrative purposes, FIGS. 5A and 5B containstwo particles, red blood cells (white dots), and capture particles(black dots). FIG. 5A illustrates blood flowing through a channelwithout a standing acoustic wave being imparted on the channel and itscontents. In FIG. 5A, the particles remain homogenously mixed throughoutthe channel. In FIG. 5B, a standing wave is imparted on the channel. Thestanding acoustic wave 501 creates two node types. A pressure nodeoccurs at 502. The node extends across the full height of the lumen. Thechannel dimensions set forth above in relation to FIG. 4A yield apressure node at approximately the center of the channel.

Particles are aligned based on the sign of their contrast factor.Particles with a positive contrast factor (e.g. the formed elements ofblood) are driven towards a pressure node 502. In contrast, particleswith a negative contrast factor (e.g. capture particles used in thesingle-stage device described above) are driven toward the pressureantinodes 503.

FIG. 6A is a top view of a separation channel 600, as depicted in FIG.2, in which fluid is flown through the separation channel 600 withoutthe application of the standing acoustic wave. The separation channel600 includes three outlets 602, 603, and 604. As with FIG. 5A, particlessuspended in the fluid are homogeneously distributed throughout thefluid, and thus are not readily discernible in the image. The particlesflow substantially evenly out of all three outlets 602, 603, and 604.

In contrast, FIG. 6B is a top view of the separation channel 600, asdepicted in FIG. 2, after the application of a standing acoustic wave,according to one illustrative embodiment.

In FIG. 6B, as a result of the standing acoustic wave, the particles 601suspended in the fluid are aligned with the middle of the separationchannel 600. Once aligned with the middle of the separation channel 600,the particles 601 exit the separation channel 600 through the middleoutlet 602. The remaining fluid, substantially devoid of particles,exits the separation channel through the side outlets 603 and 604.

FIG. 7 is an illustrative example of a lipid bilayer capture particle700. The capture particle 700 includes a lipid bilayer 701 encapsulatinga fluid 702. Anchored in the lipid bilayer are affinity particles 703.The affinity particles bind and capture target particles 704.

More specifically, the lipid bilayer 701 forms a liposomal captureparticle. In some implementations, the lipids may be, but are notlimited to, dilauroyl-glycero-phosphoglycerol anddilauroyl-glycero-phosphocholine. The capture particle is tuned foracoustically induced mobility. Entities that differ in size, density,and/or compressibility have the greatest differential mobility inacoustic fields and thus are the most readily separable. Therefore, insome implementations, the size, density, and/or compressibility of thecapture particles is modified to distinguish the capture particle fromthe formed elements of blood. The acoustic mobility of a particle isproportional to its volume. For example, in some implementations, thecapture particles are about 1 μm in diameter. In other implementations,they are between about 2 and about 5 μm in diameter. In implementationsthat adjust the compressibility of the capture particle, the rigidity ofthe capture particle can be adjusted by controlling the lipid componentsin the bilayer. The length and saturation of the lipid hydrocarbon tail,cross-linking of the hydrophobic domains, and/or the inclusion ofcholesterol can all affect the fluidity and compressibility of aliposome. In other implementations, the density of the liposome isengineered by encapsulating an acoustically active fluid 702. In theseimplementations, the acoustical active molecule can be an FDA-approvedcontrast agent, glycerin, castor oil, coconut oil, paraffin, air, and/orsilicone oil. In other implementations, all the above describedcharacteristics are manipulated to create a capture particle with thegreatest possible difference in contrast factor compared to a formedelement.

As described above, the acoustically induced mobility of a particle isbased on the contrast factor of the particle. For a liposomal basedcapture particle, the contrast factor is dominated by the properties ofthe encapsulated fluid. The contrast factor is based on the bulk modulus(K) and density (Ψ) of the encapsulated fluid. When suspended in blood,the contrast factor (⊥) for a capture particle, encapsulating a specificfluid, is calculated with the below equation:

$\begin{matrix}{\phi = {\frac{{5\rho} - {2 \cdot 1.02}}{{2\rho} + 1.02} + \frac{2.2}{K}}} & (11)\end{matrix}$

Table 4 provides the Ψ, K, and then calculated ⊥-factor based on theabove equation.

TABLE 4 Calculated Contrast Factors Materials Ψ (g/ml) K(Gpa) ⊥Encapsulated glycerin 1.25 4.7 +0.73 Fluids castor oil 1.03 2.06 −0.06coconut oil 0.92 1.75 −0.36 paraffin 0.80 1.66 0.58 silicone oil 1.041.09 −1.00 air 0.002 1.4 −3.55 Formed white blood cell 1.02 2.5 +0.12Elements red blood cell 1.10 3.0 +0.34

In some implementations, such as the implementation of FIG. 3, thecapture particles have a contrast factor that is lower in magnitude, butstill of the same sign as the formed elements. In these implementations,the low contrast factor of the capture particles can be achieved bymaking the capture particles sufficiently small to reduce their contrastfactor to below that of the formed elements.

As illustrated in FIG. 7, affinity molecules 703 are embedded in thelipid bilayer 701. In some implementations, these affinity molecules areglycoconjugates. The glycoconjugates enable the capture and retention ofall major classes of pathogens, including bacteria and viruses. In someimplementations, the affinity molecules 703 also bind to toxins andpro-inflammatory cytokines. In some implementations, affinity molecules703 are designed to universally capture gram-negative and gram-positivebacteria, viruses, and toxins, by exploiting that: 1) pathogens expressunusual surface N- and O-linked glycan structures that can be targetedby glycan-binding proteins or lectins and 2) many pathogens and toxinsbind to charged polysaccharides, especially those of the heparan sulfatefamily, that are present on the cell surface of mammalian cells. Someimplementations employ glycoconjugate capture agents that have twocomponents: a modified, non-anticoagulant heparin fragment thatnevertheless maintains high affinity, multivalent binding properties,and a glycan-binding protein that binds to surface N- and O-linkedglycans present on the surface of pathogens. In other implementations,the glycan structure is a lectin. For example the lectin can be, but isnot limited to: type 2 membrane receptors such as DC-SIGN, DC-SIGNR, andLangerine; collectins such as pulmonary surfactant proteins (SP-D,SP-Al), mannose binding lectin, and collectin-Kl; and macrophage mannosereceptors. In other implementations, the affinity molecule is anantibody.

The affinity particles are anchored to the liposomal surface so theirconcentration, valency, and distribution can be controlled. This isparticularly relevant since pathogen-receptor interactions are oftenmultivalent and the receptor configuration impacts overall avidity. Insome implementations, the affinity molecule is attached to an anchorthat is incorporated into the lipid bilayer, so the embedded functionalgroups remain in close proximity but are free to rotate and rearrange.Lipid anchors are favored because the molar ratio of derivatized lipidsincorporated can be controlled. Lectins are incorporated by solubilizinga surfactant with pre-formed liposome suspensions, through directaddition of fatty acids to lysine residues, or by modification withhydrophobic anchor lipids such as Nglutaryl-phosphotidylethanolamine(NGPE).

FIG. 8 illustrates an overview of the process of making and using acapture particle. The affinity molecules of FIG. 8A are embedded in theliposome of FIG. 8B to produce an affinity coated liposome asillustrated in FIG. 8C. Next, the capture particles are combined with ablood or other fluid containing target particles. The target particlesthen bind to the capture particles. FIG. 8E illustrates, bound targetparticles can then be removed from the fluid by acoustically moving thecapture particles whereas unbound target particles are not removed fromthe fluid.

FIG. 9 is a flow chart of a method for cleansing blood with asingle-stage microfluidic separation channel (1000). First, whole bloodis collected (step 1001). Then whole blood is flowed into an inlet of asingle-stage microfluidic separation channel, as depicted in FIG. 2(step 1002). Next, a plurality of capture particles is introduced intothe whole blood (step 1003). Then a standing acoustic wave is applied tothe separation channel (step 1004). The formed elements are thencollected in a first outlet (step 1005). Next, the capture particles arecollected in a second outlet (step 1006). Finally, the cleansed blood isreturned to a storage container or returned directly to the patient(step 1007).

Referring to FIGS. 1, 2, and 10, the method 1000 of cleansing blood witha single-stage microfluidic separation channel 200 begins by collectingwhole blood. In some implementations, the whole blood is collected froma patient 101, and then directly introduced into the blood cleansingsystem 100. In other implementations, the whole blood is collected froma patient 101 and then stored for later cleansing.

The method 1000 of cleansing blood with a single-stage microfluidicseparation channel 200 continues by flowing whole blood into the inletof a microfluidic separation channel (step 1002). The whole bloodcontains a plurality of formed elements, plasma, and a plurality oftarget particles. In some implementations, the target particles can betoxins, bacteria, and/or viruses. In some implementations, a singlemicrofluidic separation channel is used, while in others a plurality ofsingle-stage separation channels is used in conjunction to accommodategreater blood flow throughput.

The method 1000 can include introducing a plurality of capture particlesinto the whole blood (step 1003). In some implementations, theconstituent components of capture particles are injected into aseparation channel with a micronozzle and spontaneously form captureparticles as injected into the separation channel. In otherimplementations, the capture particles are prefabricated and thenintroduced into the whole blood. In some implementations, the captureparticles are introduced into the whole blood after the whole bloodenters the separation channel through the first inlet 202. In yet otherimplementations, the capture particles are introduced into the wholeblood before the blood enters through the first inlet 202 of theseparation channel 200. In some implementations, the capture particlesare microbeads and/or lipid based liposomes.

The method 1000 includes applying a standing acoustic wave to theseparation channel (step 1004). The standing acoustic wave is appliedtransverse to a direction of flow of the whole blood through theseparation channel 200. In some implementations, the formed elements andcapture particles have contrast factors with different signs. Thus, theapplication of the standing acoustic wave causes the formed elements toaggregate about the central axis of the separation channel and thecapture particles to aggregate along at least one wall of the separationchannel, as depicted in FIG. 2. In other implementations, the standingacoustic wave causes the formed elements to aggregate along at least onewall of the separation channel and the capture particles to aggregateabout the central axis of the separation channel.

The method 1000 can include collecting the formed elements of the wholeblood in a first outlet (step 1005). In some implementations, asdepicted in FIG. 2, a first outlet 204 is aligned with the central axisof the separation channel allowing the outlet to collect the formedelements as they aggregate and flow down the central axis of theseparation channel. Similarly, the method continues with the collectingof the capture particles in a second outlet (step 1006). In someimplementations, the end of the separation channel has at least a secondoutlet channel 206 and 207 aligned with at least one wall of theseparation channel. As the capture particles are driven towards theantipressure notes along the walls of the separation channel, they arecollected by the outlets channels 206 and 207 aligned with the walls ofthe separation channels. In some implementations, the standing acousticwave is adjusted such that the formed particle aligns along the walls ofthe separation channel and the capture particles align with the centralaxis of the separation channel. In such an implementation, the formedelements are funneled into outlets along the wall of the separationchannel and the capture particles are funneled into an outlet alignedwith the central axis of the separation channel. In someimplementations, the outlet channels 206 and 207 terminate in individualoutlets or merge to terminate into a single outlet 205.

The method 1000 can include the reintroduction the cleansed blood into apatient 101 or storage (step 1007). In some implementations, such assystem 100, the whole blood is collected directly from a patient andthen reintroduced to the patient 101. In some implementations, thecleansed blood is reheated to body temperature before being reintroducedinto the patient 101. In other implementations, the cleansed blood iscollected in a storage container for later reintroduction into a patient101.

The plastic-based devices described herein can be used to separateacoustically active particles from fluids. The devices described hereincan be used with or without capture particles. For example, particlesand cells (e.g., target particles) can be removed from a fluid based onthe target particles' acoustic properties with respect to the fluid inwhich they are contained. The fluids can be biological based (e.g., abodily fluid such as blood) or non-biological based (e.g., waste water).In one example, the plastic-based devices described herein can be usedin the isolation of natural blood cells such as hematopoietic stem cellsor T-lymphocytes. The device can be used in cell therapy andbioprocessing. For example, the device can separate out hematopoieticstem cells or T-lymphocytes based on their acoustic activity withrespect to the fluid (e.g., blood) flowing through the device. Thehematopoietic stem cells or T-lymphocytes can be driven to anaggregation axis at a rate different than the other elements in thefluid enabling the hematopoietic stem cells or T-lymphocytes to beseparated from the fluid at different points along the length of thedevice. In other implementations, debris or other target particles canbe removed from fluid flowing through the device. For example, theparticles or debris can be removed from large scale batches of culturedcells (e.g. mesenchymal stem cells) prior to therapeutic injection ofsuch cells.

In some implementations, the plastic-based devices described herein canbe used in antibiotic susceptibility testing. For example, the devicecan be a component of a system for bacterial identification that caninclude a single, self-contained, point-of-care or lab-based diagnosticsystem. The system can be used to detect foreign agents, such asbacteria, within blood or other samples. The system can receive as inputthe blood or other samples and output an indication of whether, and towhat degree, the foreign agent is present in the sample. The system canreduce the time scale for bacteria detection to a few hours and serve asa point-of-care diagnostic tool within hospital, lab, and other medicalfacilities. The system can include disposable microfluidic cartridgesthat include the plastic-based separation device that are removable fromthe system and can be replaced between tests. The microfluidiccartridges can receive a sample, such as a blood sample, that issuspected of containing bacterial cells and separate the bacterial cellsfrom the blood sample. Once the bacterial cells are separated from theblood, the system can introduce recombinant detector phages (RDBs) intothe system. The RDB can include one or more reporter genes. When the RDBcomes into contact with a specified bacterial cell type, the RDB caninfect the bacterial cells with the reporter gene. Once infected, thebacterial cells can then express the reporter gene. The system candetect a signal generated responsive to the expression of the reportergene with an optical detector. The signal can include luminescence,fluorescence, or chromagraphic signals generated in response to theexpressed reporter gene. The system can display or otherwise report outthe signal as an indication of the presence of the foreign agent.Additional information of a bacterial identification system can be foundin U.S. patent application Ser. No. 15/470,750, which is incorporated byreference in its entirety.

EXAMPLES

Different separation channels were constructed in thermoplastics to testand illustrate the improved performance of devices designed using thetechniques described herein, such as the method 450. The performance ofthe channels using the herein described methods was measured using theprominence of local maxima. Prominence can have the advantage over rawpixel intensity for the purposes of comparison due to itsself-normalizing nature. Since prominence is measured relative to pointson the signal itself it is robust against irregularities inherent to thesignal. These irregularities can take the form of variable lightingconditions between experimental runs, such as variations inenvironmental lighting, and illumination variabilities within a singlemicroscope image's region of interest, such as skewed backgroundintensities caused by shadows.

Peak prominence is used as a direct measure of merit for devicescreening studies in which the assumption of a functional geometry(e.g., a geometry capable of focusing particles) cannot be made. When adevice is determined to function well based on its prominence score itis compared to the baseline geometry using the ratio of peak prominenceto half-prominence width, χ. χ cannot be used during device screening asthe metric can skew results by rewarding peaks with relatively smallprominence values and correspondingly small widths. However, this metricis useful for comparing the quality of the most prominent peaks amongdifferent, commensurate, devices such as the winner of a designscreening iteration and the baseline geometry.

A device's performance can be tested head-to-head against the baselinegeometry using the full, trifurcated, designs shown in FIG. 2. Theperformance testing can include comparing each design's ability to focusblood to the center channel, as shown in FIG. 2, as well as eachdesign's ability to separate bacteria from blood.

The half-prominence width of a peak of prominence Prom is calculated bydrawing two horizontal lines extending in the negative and positivedirections from the point of half-prominence. These lines extend ineither direction until either the end of the signal is reached or theline intersects the signal itself. The indices of these events in thenegative and positive directions are recorded as i⁻ and i⁺,respectively. The peak width Half Prom Width is then defined as |i⁺−i⁻|.The final equation for χ:

$\begin{matrix}{\chi = {{Prom}*\frac{W_{c}}{{Half}\mspace{14mu} {Prom}\mspace{14mu} {Width}}}} & (12)\end{matrix}$

Constructing a separation channel using the above formulas for athermoplastic device, a separation channel with a channel width of 550μm, a channel height of 250 μm, a side-wall width of 850 μm wasconstructed (and is referred to as the experimental design). Thisexperimental device was compared against a baseline device using thedimensions as illustrated in FIG. 3. The comparison of the performanceof the separation device is illustrates in FIGS. 10A-10C. The solid lineillustrates the normalized prominence vs average dissipated power (mW)of the experimental device constructed using the equations for athermoplastic device and the dashed line illustrates the normalizedprominence of the baseline device. FIG. 10A illustrates the operation ofthe devices with a 25 μl/min flow rate, FIG. 10B with a 50 μl/min flowrate, and FIG. 10C with a 75 μl/min flow rate. As illustrated in FIGS.10A-10C, the normalized prominence (indicating a grayscale/pixel) isgreater at each average dissipated power level and at each of the testedflow rates for the experimental device than for the baseline device. Theprominence can indicate the ability of the device to concentrateparticles near the middle of the separation channel. Accordingly, FIGS.10A-10C illustrate that the experimental device was better able to focusparticles near the middle of the separation channel when compared to thebaseline device.

FIGS. 11A-11C illustrate plots of the performance of the experimentaldevice against the baseline in terms of each device's ability to focus(e.g., divert) red blood cells (RBC) to a central aggregation axis. Thedynamic range of each measurement was limited to that which fallsbetween the performance at the control measurement (e.g., zero averagedissipated power) and 100% RBC concentration in the center channel. Asthe chip design used, shown in FIG. 2, has a single input port and twooutlet ports, RBCs will be equally distributed between the two outletports in the acoustics-off (0 W input power) condition.

In FIGS. 11A-11C the solid lines indicate the RBC concentration exitingthe center channel of the experimental device at each of a plurality ofdifferent dissipation powers. The dotted lines indicate the RBCconcentration exiting the center channel of a baseline device at each ofa plurality of different dissipation powers. The baseline andexperimental design demonstrated comparable performance at a flow rateof 25 μl/min across all power settings (FIG. 11A); however, at higherflow rates (e.g., 50 μl/min in FIG. 11B and 75 μl/min in μl/min FIG.11C) the experimental design outperformed the baseline across allnon-control power settings as illustrated by the experimental device'scentral channel having a higher RBC concentration.

Additionally, four experiments were conducted, two for each devicedesign (baseline and experimental), in order to determine the optimalvalue for each measure of merit while holding the other constant.Optimality was defined as the maximum flow rate or minimum powerrequired to maintain 90% RBC separation between the side and centerports while achieving equivalent bacteria-blood separation performance.

FIG. 12A illustrates the relative performance of the baseline device asit compares to the experimental design in terms of flow rate with powerheld constant at 1 W. Each of the devices are performing comparableseparation of bacterial cells from blood. FIG. 12B illustrates theperformance of the two chip designs holding flow rate at 50 μL/min whilevarying power. Each of the devices are performing comparable separationof bacterial cells from blood. Constant values were maintained for RBCseparation and bacterial recovery for all chip designs and experimentsFIG. 12A illustrates that the experimental design achieved a 175%increase in throughput relative to the baseline design. Additionally,the experimental design was able to decrease the average dissipatedpower by 81.63% when compared to that of the baseline geometry. Theactual average RBC separation across all four experiments was 95.25%(±1.89%). The purity of the bacterial samples collected at the side porthad a standard deviation of 0.05%. Additionally, the experimental designachieved better RBC separation and equivalent bacterial recoveryrelative to the baseline for both the maximum flow rate and minimumpower experiments (98% vs 94% and 95% vs 94%, respectively).

What is claimed:
 1. A device, comprising: a separation channel definedwithin at least one thermoplastic substrate, the separation channelhaving a width, a first wall, a second wall, a roof, and a floor,wherein the floor is configured to couple with an acoustic transducerand a thickness of the first wall and the second wall is determined bythe width of the separation channel; a first inlet defined within the atleast one thermoplastic substrate to introduce a fluid into a proximalend portion of the separation channel; a first outlet defined within theat least one thermoplastic substrate, the first outlet positioned at adownstream portion of the separation channel substantially along thelongitudinal axis of the separation channel; and a second outlet definedwithin the at least one thermoplastic substrate, the second outletpositioned at the downstream portion positioned adjacent to a first wallof the separation channel.
 2. The device of claim 1, wherein thethickness of the floor is determined by the width of the separationchannel.
 3. The device of claim 1, wherein the width of the separationchannel is between 0.1 mm and 3 mm.
 4. The device of claim 1, wherein awavelength of an acoustic wave generated by the acoustic transducer isbetween about 3.3 and about 5 times the width of the separation channel.5. The device of claim 1, wherein the thickness of the first wall andsecond wall is based on a velocity of an acoustic wave through thefluid.
 6. The device of claim 5, wherein the thickness of the first walland the second wall is based on a velocity of the acoustic wave throughthe first thermoplastic substrate.
 7. The device of claim 1, wherein theheight of the separation channel is determined by the width of theseparation channel.
 8. The device of claim 1, wherein a ratio of theseparation channel to the height of the separation channel is between 2and 2.5.
 9. The device of claim 1, wherein the floor of the separationchannel has a first thickness and the roof of the separation channel hasa second thickness.
 10. The device of claim 9, wherein the firstthickness is greater than the second thickness.
 11. The device of claim1, wherein a ratio of the thickness of the floor of the separationchannel to a width of the separation channel is between about 0.5 and 1.12. The device of claim 1, further comprising a plurality of separationchannels defined within the at least one thermoplastic substrate, eachof the plurality of separation channels separated by an air gap.
 13. Amethod to process fluid, comprising: providing a separation devicecomprising: a separation channel defined within at least onethermoplastic substrate, the separation channel having a width, a firstwall, a second wall, a roof, and a floor, wherein the floor isconfigured to couple with an acoustic transducer and a thickness of thefirst wall and the second wall is determined by the width of theseparation channel; a first inlet defined within the at least onethermoplastic substrate to introduce a fluid into a proximal end portionof the separation channel; a first outlet defined within the at leastone thermoplastic substrate, the first outlet positioned at a downstreamportion of the separation channel substantially along the longitudinalaxis of the separation channel; and a second outlet defined within theat least one thermoplastic substrate, the second outlet positioned atthe downstream portion positioned adjacent to a first wall of theseparation channel; flowing a fluid through the first inlet, wherein thefluid comprises undesirable particles; driving, with a standing acousticwave generated by the acoustic transducer, the undesirable particlestoward the first wall of the separation channel.
 14. The method of claim13, wherein the width of the separation channel is between 0.1 mm and 3mm.
 15. The method of claim 13, wherein the width of the first wall andsecond wall is based on a velocity of the acoustic wave through thefluid and a velocity of the acoustic wave through the first wall and thesecond wall.
 16. The method of claim 15, wherein the width of the firstwall and the second wall is based on a velocity of the acoustic wavethrough the first thermoplastic substrate.
 17. The method of claim 13,wherein the height of the separation channel is based on the width ofthe separation channel.
 18. The method of claim 13, wherein a ratio ofthe separation channel to the height of the separation channel isbetween 2 and 2.5.
 19. The method of claim 13, wherein the floor of theseparation channel has a first thickness and the roof of the separationchannel has a second thickness and the first thickness is greater thanthe second thickness.
 20. The method of claim 13, wherein a ratio of thethickness of the floor of the separation channel to a width of theseparation channel is between about 0.5 and 1.